Transcript Engineering Gut Microbiota

ENGINEERING GUT MICROBIOTA
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Fei Chen
5/11/08
THE BIG PICTURE
Gut microbiota provide a dynamic and very
beneficial symbiotic relationship with human
hosts.
 Gut microbiota perform key functions in
metabolism
 They influence drug response, and the
development of many diseases.
 Genetically engineered gut bacteria can have farranging impact on health and quality of life.
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SOME BACKGROUND
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Present in immense numbers of in the gut:~1 x 1014
bacteria.
30-40 species compose of 99% of the population.
Mutualistic/symbiotic relationship with hosts. In humans,
they serve several key roles:
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Very significant for health:
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Metabolism and fermenting unused energy substrates.
Training the immune system
Production of vitamins.
And competitive inhibition of harmful species.
Obesity
Disease
Aging
‘Gut’ refers to the general digestive tract, in terms of
microflora, we are interested in the large intestine,
especially the Cecum, where a large population of bacteria
is present.
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THE QUESTION POSED IS:
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How can we engineer bacteria as to make a
superior gut microbe?
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IS IT VIABLE? SOME CONSIDERATIONS…
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Gut flora maintains a dynamic relationship with
the host immune-system. The introduced
engineered bacteria must not promote a immuneresponse from the host.
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Research has already shown that a specific strain of
E. Coli, NISSLE 1917, can be engineered and
introduced to the mice gut without provoking an
immune response.
Gut flora is very sensitive to environmental
changes. Small changes in pH or population can
cause drastic changes to population make-up.
 We have to consider the interaction between our
genetically engineered flora and the local
microbes.
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POSSIBLE IDEAS
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There are many possible applications in
engineering gut microbes. Five projects based on
these ideas are:
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Gut pH management
Vitamin Production
Lactase Production/Digestion
Pathogenic Defense
Varied expression through Slipped Strand
Mismatching
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GUT PH MAINTENANCE
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The flora in the gut is very sensitive to pH.
The body maintains a healthy bacterial ecosystem
using pH. Our bacteria could do the same.
 The pH of an healthy adult Cecum is around 6.4. (std.
dev. 0.4)
 Design a system which maintains the pH in this
range.
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System Requirements:
Sense and respond to external pH.
 Create byproducts which buffers external pH.
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Reducing pH would be easy, acids are byproducts of
metabolism.
 Increasing pH is harder.
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VITAMIN PRODUCTION
Bacteria are responsible for the production of
several vitamins required for the body. Example:
E. Coli produces vitamin K.
 Engineer bacteria which produce other Vitamins
and nutritional benefits.
 One example: Beta-Carotene.
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Lycopene cyclase, an enzyme separated from
cyanobacteria which produces Beta-Carotene from
lycopene.
 Lycopene is commonly found in human diet, a source
is tomatoes.
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Design Goal: Engineer bacteria which can
produce Beta-Carotene/ Vitamins.
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LACTASE PRODUCTION AND DIETARY
REGULATION
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Lactose is a sugar predominantly found in milk.
Lactose intolerance is the inability to metabolize
glucose, and is found in a large percentage of Asians
and Africans. It is estimated that 70% of adults are
lactose intolerant.
Lactase is a glycoside hydrolase which breaks lactose
disaccharides into galactose and glucose monomers.
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This gene can be incorporated into our engineered bacteria
to metabolize lactose in those who have lactose intolerance.
Further ideas in this subsystem include dietary
regulation.
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Bacteria has been shown to have a significant link with
obesity. The cause of this is due to the balance between
various subpopulations inhabiting the gut. (How to exploit
this is unclear)
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FEND OFF PATHOGENS
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Example Pathogen:
Salmonella (S.
Typhimurium )
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One of the major causes of
food poisoning. Found in
inadequately cooked eggs,
pets.
Release of lysogenic phage
held in our engineered
bacteria to infect and lyse
salmonella.
P22 is an well documented phage which infects
Salmonella, and has lysogenic/lytic life-cyles. Can
enter lytic life cycle after UV-exposure/DNA
damage.
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OVERALL SYSTEM CONSTRUCTION
Use of Slipped-Strand-Mismatching to generate
different subpopulations with varied expression.
 Possible Ways to Utilize the Idea:
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Combinations of Slip-Strand Mismatching devices to
create many different phenotypes. We have multiple
functions needed by the cell.
 Set one phenotype to be more predominant than
others. (Example: Set Vitamin production to be the
primary phenotype)
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PROS AND CONS
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Many different subsystems which can be constructed
because gut microbiota do so much.
Bacteria already inhabit the gut, including as our
favorite, E. Coli.
Many of the subsystems are very interesting by
themselves and could be very useful for future teams:
Bacterial Phage Production
 Slipped-Strand Mismatching for multiple subpopulations.
 External pH buffering
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The flora of the gut is very complicated and very
sensitive.
A lot of work required for each subsection.
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REFERENCES:
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Controlling the metabolic flux through the carotenoid
pathway using directed mRNA processing and
stabilization. C D Smolke, V J Martin, and J D Keasling
Metab Eng. 2001 October; 3(4): 313–321.
Microbial ecology: Human gut microbes associated with
obesity Ruth E. Ley, Peter J. Turnbaugh Nature 444, 10221023.
Measurement of gastrointestinal pH profiles in normal
ambulant human subjects. D F Evans, G Pye, R Bramley, A
G Clark, T J Dyson, and J D Hardcastle Gut. 1988 August;
29(8): 1035–1041.
Intestinal immunity of Escherichia coli NISSLE 1917: a
safe carrier for therapeutic molecules Astrid M.
Westendorf, Florian Gunzer, Stefanie Deppenmeier FEMS
Immunol Med Microbiol 2005 Mar 1; 43(3) 373-84.
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